A. Tucholska

Influence of long-term equine herpesvirus type 1 (EHV-1) infection on primary murine neurons—the possible effects of the multiple passages of EHV-1 on its neurovirulence

Equine herpesvirus 1 (EHV-1), like other members of the Alphaherpesvirinae subfamily, is a neurotropic virus causing latent infections in the nervous system of the natural host. In the present study, we have investigated EHV-1 replication (wild-type Jan-E strain and Rac-H laboratory strain) during long-term infection and during the passages of the virus in cultured neurons. The studies were performed on primary murine neurons, which are an excellent in vitro model for studying neurotropism and neurovirulence of EHV-1. Using real-time cell growth analysis, we have demonstrated for the first time that primary murine neurons are able to survive long-term EHV-1 infection. Positive results of real-time PCR test indicated a high level of virus DNA in cultured neurons, and during long-term infection, these neurons were still able to transmit the virus to the other cells. We also compared the neurovirulence of Rac-H and Jan-E EHV-1 strains after multiple passages of these strains in neuron cell culture. The results showed that multiple passages of EHV-1 in neurons lead to the inhibition of viral replication as early as in the third passage. Interestingly, the inhibition of the EHV-1 replication occurred exclusively in neurons, because the equine dermal (ED) cells co-cultivated with neuroculture medium from the third passage showed the presence of large amount of viral DNA. In conclusion, our results showed that certain balance between EHV-1 and neurons has been established during in vitro infection allowing neurons to survive long-term infection.

Acyclovir and trichostatin A modulate EHV-1 replication in murine neurons in vitro

Equine herpesvirus type 1 (EHV-1) is one of the most important viral pathogens of horses worldwide (2). It may cause respiratory disease, sporadic or epizootic abortions, or, recently more often, neurological disease known as equine herpesvirus myeloencephalopathy (EHM), which may be life-threatening and results in significant economic losses to the equine industry (1, 13, 15, 18, 19). It has been suggested that a single-nucleotide polymorphism in the EHV-1 DNA polymerase gene, which leads to amino acid variation (N752/D752), may be associated with outbreaks of EHM (3, 11, 13). D752 strains of EHV-1, which are statistically more often isolated from cases of EHM, were called neuropathogenic strains. However, it is worth mentioning that all EHV-1 strains show neurotropism and are capable of establishing latency in peripheral neurons. Moreover, EHV-1 may also establish latency in leukocytes (10). The main role of latency is to maintain the viral genome for a long time inside host cells, at the same time avoiding the immune response. On the other hand, the virus may reactivate and start productive replication at any time, especially during stress, which leads to the dissemination of progeny virions (17). The current approach to the control of EHV-1 infections is based on biosecurity measures and vaccination, but it is not sufficient. Immunity after infection or vaccination is usually incomplete and short-lived, and once latency has been established, the virus cannot be eliminated from host cells. Although some progress has been made in understanding the adaptive immunity to EHV-1, innate immunity remains poorly characterized, despite the fact that it is critically important for inducAcyclovir and trichostatin A modulate EHV-1 replication in murine neurons in vitro1)